A typical lateral flow test utilizes the concept of lateral liquid or suspension flow in order to transport a given sample to the test. These types of tests are used for a wide variety of applications, including diagnostics (e.g., pregnancy and other types of medical testing) and environmental testing.
Typically, a lateral flow test may require as many as five separate materials in order to optimize the test. The materials serve as a wick to transport the sample to the test; as a filtration material to remove unwanted particles; as a conjugate release pad where the detection reagent(s) is immobile when dry but mobilized when wet; as a reaction matrix where the capture reagents are immobilized; and as an absorbent where the sample is absorbed and the liquid is driven to flow along the test format.
Despite the wide array of usages, lateral flow tests are frequently subject to flow problems and are complicated to manufacture. These tests are complex, multipart assays performed on a series of overlapping pads of different types of materials aligned on a test strip. Problems arise from material incompatibility, contact issues, and imperfect material characteristics. Boundaries found between segments can adversely affect flow characteristics. Different materials may have widely different flow, or wicking, rates and may have different effects on molecules flowing through them.
Currently, different materials are used for each part of the test, due to the vastly different physical characteristics needed for each component. For example, the sample wick must be fast wicking and have a very open structure; the filtration material must have a pore size of the correct size to remove the unwanted particles; the conjugate release must be non-protein binding; the reaction matrix must be protein binding and consistent. Due to the different properties required, it is normal for a test to be made up of overlapping pads of several different materials. Generally, a membrane, such as a nitrocellulose membrane, is used for the reaction matrix; glass fiber or man-made fibers (e.g., cellulose) are used for the sample application/filtration layer and for the conjugate release layer; and cellulose or glass fiber materials are used for the absorbent (Whatman plc).
Typically, a sample is placed on a sample application wick (e.g., glass fiber, cast cellulose acetate, fused PE, or cellulose fiber), where the wicking process begins. Optionally, the sample runs through the wick and into and through a filtration pad (e.g., glass fiber, glass membrane, cellulose fiber, cast cellulose acetate, fused PE, man-made fibers, and mixtures of man-made fibers and glass fibers), which may be used to remove contaminants or, for example, to remove erythrocytes (red blood cells) in a blood sample in order to eliminate them from the sample or to prevent their red coloration from interfering with a downstream color indicator. Next, the sample wicks into a conjugate pad (e.g., glass fiber or polyester), where the sample liquid or suspension mixes with the colored conjugate reagent (e.g., an antibody), causing the conjugate reagent to be released. If the sample is positive, the conjugate will bind to the analyte. Both bound and unbound conjugate will flow laterally through the conjugate pad into capture area pad, which is typically nitrocellulose. In some examples, the capture area pad may comprise two lines of protein striped perpendicularly onto the nitrocellulose membrane. One line (test) binds to the analyte (if present), while the other (control) binds to the conjugate in order to indicate that the test itself has been successful, regardless of positive or negative result. Therefore, a successful positive test shows two lines (test and control), while a successful negative test shows one line (control only). The absorbent pad, which is typically cellulose or glass fiber, acts as an absorbent to pull the liquid through the strip. The entire assembly of overlapping pads, each having one or more layers, is attached to an assembly sheet, which may be made of various types of materials (e.g., plastic) and which does not interact with the test.
It would be desirable to have a single-layer lateral flow format to reduce flow problems due to material incompatibility and contact issues, to decrease development time, to improve accuracy and efficiency of lateral flow test results, to provide superior performance, to lower manufacturing costs, and to aid in the ease of use of the format.